We report a case of Ancylostoma ceylanicum infection detected by endoscopy. It was diagnosed and confirmed using polymerase chain reaction (PCR) and DNA sequencing. The patient is a 58-year-old Malaysian woman who lives in a rural area, where uncontrolled populations of stray and semidomesticated dogs live in close proximity with humans.
Currently, information on species-specific hookworm infection is unavailable in Malaysia and is restricted worldwide due to limited application of molecular diagnostic tools. Given the importance of accurate identification of hookworms, this study was conducted as part of an ongoing molecular epidemiological investigation aimed at providing the first documented data on species-specific hookworm infection, associated risk factors and the role of domestic animals as reservoirs for hookworm infections in endemic communities of Malaysia.
Ancylostoma ceylanicum is a common zoonotic nematode. Cats act as natural reservoirs of the hookworm and are involved in transmitting infection to humans, thus posing a potential risk to public health. The prevalence of feline A. ceylanicum in Guangzhou (South China) was surveyed by polymerase chain reaction-restriction fragment length polymorphism (PCR-RFLP). In total, 112 faecal samples were examined; 34.8% (39/112) and 43.8% (49/112) samples were positive with hookworms by microscopy and PCR method, respectively. Among them, 40.8% of samples harboured A. ceylanicum. Twelve positive A. ceylanicum samples were selected randomly and used for cox 1 sequence analysis. Sequencing results revealed that they had 97-99% similarity with A. ceylanicum cox 1 gene sequences deposited in GenBank. A phylogenetic tree showed that A. ceylanicum isolates were divided into two groups: one comprising four isolates from Guangzhou (South China), and the other comprising those from Malaysia, Cambodia and Guangzhou. In the latter group, all A. ceylanicum isolates from Guangzhou were clustered into a minor group again. The results indicate that the high prevalence of A. ceylanicum in stray cats in South China poses a potential risk of hookworm transmission from pet cats to humans, and that A. ceylanicum may be a species complex worldwide.
Soil-transmitted helminth infections remain a major public health burden in low- and middle-income countries. The traditional diagnosis by microscopic examination of fecal samples is insensitive and time-consuming. In this study, a pentaplex real-time polymerase chain reaction (PCR) was evaluated for the simultaneous detection of Ancylostoma, Necator americanus, Ascaris lumbricoides, and Strongyloides stercoralis. The results were compared with those obtained by conventional parasitological diagnostic methods. Real-time PCR was positive in 48 of 77 samples (62.3%) and microscopic examination was positive in six samples (7.8%) only (P < 0.05). In conclusion, the real-time PCR assay described in this study provides a specific and sensitive diagnostic tool for the detection of these four helminth species in epidemiological studies and monitoring of treatment programs.
Ancylostoma ceylanicum is the only zoonotic hookworm species that is able to produce patent infections in humans with the majority of cases reported in South East Asia. Over the past few years, there have been an increasing number of studies investigating the prevalence of this parasitic zoonosis using molecular diagnostic tools and a single genetic locus as marker for species identification. As there can be limitations in using a single genetic locus for epidemiological studies and genetic discrimination, the complementary use of a more variable locus will provide additional evidence to support the zoonotic exchange of hookworm species between humans and animals. In the present study, the cytochrome c oxidase subunit 1 (cox 1) sequence of A. ceylanicum from positive human and animal fecal samples were determined and compared with published reference sequences. Phylogenetic analysis demonstrated that isolates of A. ceylanicum were divided into two clusters, one consisting 3 human isolates and the other comprising 19 isolates of human and animal origin from different geographical locations within Malaysia. The two groups of A. ceylanicum could be distinguished from one another through five fixed nucleotide differences at locations 891, 966, 1008, 1077 and 1083. The detection of genetically distinct groups and considerable level of genetic variation within the cox 1 sequence of A. ceylanicum might suggest potential haplotype-linked differences in zoonotic, epidemiological and pathobiological characteristics, a hypothesis that still needs further investigation.
BACKGROUND: Hookworm infection is one of the important Neglected Tropical Diseases (NTD) in the world. It was previously more prevalent in the northern and southern parts of Iran with a prevalence rate higher than 40% in some endemic regions; nevertheless, the infection rate has decreased to less than 1%. This study aimed to determine prevalence and molecular aspects of hookworm infections in rural inhabitants of Fouman County, Guilan Province, northern Iran.
METHODS: This cross-sectional study was performed in 31 villages of Fouman district in Guilan Province, northern Iran during 2015-2016. Stool samples were collected from 1500 rural inhabitants and examined by formalin ethyl-acetate concentration as well as agar plate culture techniques. After treatment with albendazole, adult hookworms were isolated. Following DNA extraction, PCR amplification of ITS2-rDNA region was performed and the product was sequenced, followed by genetic variation analysis.
RESULTS: Of 1500 samples, one case was morphologically diagnosed as N. americanus. In addition, molecular characterization verified the presence of N. americanus, showing more than 95% similarity with sequences of N. americanus present in GenBank. The patient showed no clinical symptoms and a mild hypereosinophilia was the only laboratory finding observed.
CONCLUSION: A reduced prevalence of human hookworms was demonstrated within Guilan Province located in north of Iran. The N. americanus originated from Guilan had a high homology with the isolates found in Japan, Laos, Malaysia, and Australia.